结肠息肉一碳代谢通路关键酶异常调控的机制研究

Abstract: One-carbon metabolism pathway is the only methyl donor for epigenetic modification and DNA synthesis. Folate deficiency or aberrant key enzymes expression in this pathway will affect its function, leading to epigenetic abnormalities, and promoting colorectal carcinogenesis. However, the molecular mechanisms of one-carbon metabolism pathway in colorectal polyps are not well elucidated and folate supplementation in colorectal polyps chemoprevention is still controversial. In preliminary works, we screened the one-carbon unit donor amino acids in serum and tissue of patients with colorectal polyps and the healthy controls by UPLC-MS (Ultra Performance Liquid Chromatography tandem Mass Spectrometry). Interestingly, we found that serine, glycine, histidine and methionine increased in serum of patients compared to controls, moreover, the transcription levels of DNMT (DNA methyltransferase) and DHFR (Dihydrofolate reductase), both key enzymes of one carbon metabolism, also raised in colorectal polyps tissues compared to normal mucosa, suggesting the key enzymes might play an important role in colorectal polyps pathogenesis by regulating the one-carbon metabolism pathway. Further studies will investigate the expression level of whole key enzymes in one-carbon metabolism as well as the variation of one-carbon unit carrier, and test the key enzymes functions by modulating the enzymes expression in vitro. The study’s results will help to further understand upon the role of one-carbon pathway played in colorectal polyps pathogenesis, and scientifically guide the therapeutic of colorectal polyps.

表观遗传修饰是调控结肠息肉形成的主要机制之一。作为表观遗传调控的重要参与者和体内甲基化唯一供体，一碳代谢通路是目前生物学研究热点。叶酸摄入不足或一碳代谢关键酶表达异常均可影响该通路，导致表观遗传修饰改变。申请人前期通过先进的超效液相质谱技术，首次全面定量检测了结肠息肉患者及对照人群的血清及息肉组织中，该通路的一碳单位供体氨基酸水平，发现患者血清和组织中的多种供体氨基酸水平（丝氨酸、甘氨酸及甲硫氨酸）较对照组显著增高；并发现息肉组织中一碳代谢关键酶DNMT、DHFR转录水平亦较对照组显著升高。提示一碳代谢关键酶可能通过调控该通路，促进结肠息肉的形成。进一步拟在血清和组织水平全面建立结肠息肉中一碳代谢通路各种分子的表达谱改变，并在体内外探讨调控关键酶基因的表达对结肠息肉发生发展的意义。预期成果将有助于明确一碳代谢通路在结肠息肉发病中的作用，并为结肠息肉的干预提供科学依据，具有重要价值。

肠息肉为慢性炎症引起局部粘膜增生肥厚而形成的隆起样病变，肠息肉是结肠癌高危因素，腺瘤性息肉85%会演变成腺癌。因此研究息肉的形成机制与治疗方式，具有重大意义。此项研究结果包括机制研究及治疗研究。. 肠息肉机制研究发现：一碳代谢通路作为表观遗传调控的重要参与者和体内甲基化唯一供体，通过调控表观修饰参与肠息肉的发生。1）息肉患者血清一碳单位供体氨基酸水平明显高于正常人群。2）通过应用UPLC-MS发现息肉旁粘膜的氨基酸（丝氨酸、甘氨酸、甲硫氨酸、半胱氨酸等）含量较正常人群显著增高，而息肉组织中氨基酸含量较息肉旁粘膜及正常人群显著降低；3）息肉组织的氨基酸转运体水平亦显著增高，或可说明息肉氨基酸水平降低由其消耗所致。4）息肉组织整体甲基化及羟甲基化水平明显低于息肉旁和正常人粘膜；5）息肉组织中叶酸及其代谢物水平（叶酸、四氢叶酸、5-甲基四氢叶酸、5-10亚甲基四氢叶酸和5-10亚甲酰四氢叶酸水平）明显低于正常人肠粘膜和息肉旁粘膜。推论：一碳代谢通路及氨基酸代谢异常是肠息肉发生的潜在促进因素，因此通过干预关键氨基酸或叶酸可能成为预防息肉的重要措施。. 肠息肉治疗研究发现：1）首次在体外成功构建息肉类器官培养系统，该系统具有可传代、增殖快、可重复性好等优点，该系统可望成为有潜力的药物筛选平台，弥补了肠息肉缺少稳定细胞实验研究的空缺；2）依托该系统筛选多种治疗药物，并成功发现二甲双胍可显著抑制息肉类器官增殖，并初步阐明该作用的分子机制，即二甲双胍通过干预糖酵解发挥抑制功能。肠息肉类器官培养系统的构建，弥补了基础实验模型的空缺，并可作为开发息肉治疗药物的良好筛药工具。